ABSTRACT

Nonalcoholic fatty liver disease (NAFLD) is a common complication of obesity and type 2 diabetes mellitus (T2DM). Most times it is an unrecognized comorbidity to the primary care provider and endocrinologist. Today it is the most common chronic liver disease in developed countries. It is characterized by insulin resistance and hepatic triglyceride accumulation in the absence of co-existing etiologies, such as excessive alcohol consumption, viral hepatitis, medications or other etiologies for hepatic steatosis. Its more severe form of the disease with steatohepatitis (NASH) is associated with hepatocyte injury (necrosis and inflammation) and frequently with fibrosis. Although it appears to be an indolent condition, with few symptoms and often normal plasma aminotransferases, NASH is a leading cause of end-stage liver disease and hepatocellular carcinoma (HCC), and significantly increases the risk of developing cardiovascular disease (CVD) and T2DM. The pathogenesis of NASH remains poorly understood, and likely to be multifactorial, but insulin-resistant adipose tissue plays an important role. The natural history of NAFLD is incompletely understood, but risk factors for disease progression include weight gain, obesity and T2DM, as well as the severity of fibrosis stage at diagnosis. Diagnostic algorithms are evolving but we offer an approach that integrates for the non-hepatologist plasma biomarkers, imaging, and the role of liver biopsy for the management of these complex patients. At the present time, early screening -with biomarker panels or a liver ultrasound, ideally with transient elastography- is reserved for high-risk patients (i.e., obese patients with T2DM or elevated plasma AST/ALT levels or evidence of steatosis at a random liver exam) until more accurate non-invasive methods are available. A liver biopsy should be considered on a case-by-case basis, to identify those at risk of NASH-cirrhosis, working in close collaboration with a hepatologist. Treatment should include a comprehensive approach with lifestyle modification and therapeutic agents tested in RCTs, such as vitamin E (in patients without diabetes) or pioglitazone for patients with or without diabetes. Pioglitazone, given its low-cost as a generic medication, long-standing track record of efficacy in NASH, and cardiometabolic benefits, is likely to be for NASH what metformin has become for the management of T2DM. However, proper patient selection and close monitoring is needed. In addition, a number of new pharmacological agents are being studied in phase II/III trials and future management will involve the use of combination therapy, as for other chronic metabolic conditions. In summary, endocrinologists need to be aware of the severe metabolic and liver-specific complications of NASH and establish early-on a long-term management plan. Screening will likely take place in the same way as for diabetic retinopathy or nephropathy. A better understanding of its natural history and pathogenesis of NASH, combined with improved diagnostic and treatment options, will likely place endocrinologists at the forefront of the management efforts to prevent end-stage liver disease in patients with NASH. For complete coverage of all related areas of Endocrinology, please visit our on-line FREE web-text, WWW.ENDOTEXT.ORG.

INTRODUCTION

Nonalcoholic fatty liver disease (NAFLD) is a chronic liver condition that is on the rise. It has become the most common chronic liver condition in many parts of the world. It encompasses a wide spectrum of disease with different clinical implications. NAFLD means that there is evidence of liver steatosis, either by imaging or histology, in the absence of secondary causes of hepatic fat accumulation such as significant alcohol consumption, chronic use of steatogenic medication, or another established chronic liver disease. Between 40 to 50% of patients that are obese have NAFLD and this rises to about 70% if they have type 2 diabetes mellitus (T2DM). In its simplest form, known as isolated steatosis or NAFL (nonalcoholic fatty liver), there is triglyceride accumulation of ≥5% without evidence of hepatocellular injury in the form of hepatocyte ballooning or evidence of fibrosis. Although the natural history of this condition remains uncertain, and may possibly progress to more severe disease, at the present moment NAFL is considered to be associated with limited risk of liver morbidity. However, it is associated with insulin resistance so that the liver can be seen as a “mirror” of metabolic health (i.e., in obesity steatosis being a reflection of insulin resistance, and in particular, of adipose tissue dysfunction) and with an increased risk of cardiovascular disease (CVD). In its more severe form, known as nonalcoholic steatohepatitis or NASH, steatosis ≥5% is associated with hepatocyte injury with necrosis (“ballooning”) and lobular inflammation, with or without fibrosis.

Steatohepatitis is often a progressive disorder in T2DM associated with the development of fibrosis that can eventually lead to cirrhosis. Liver fibrosis is defined by its severity in stages ranging from absence of fibrosis (stage F0) to mild (stage F1), moderate (stage F2, with zone 3 sinusoidal fibrosis plus periportal fibrosis), and “advanced” fibrosis referring specifically to stages 3 (bridging fibrosis) or 4 (cirrhosis). Having fibrosis is the most important histological feature of NAFLD associated with long-term mortality. Fibrosis also predisposes patients to hepatocellular carcinoma (HCC).

Type 2 diabetes mellitus has been a well-established factor in the progression of NAFLD to more severe forms, including a higher incidence of HCC (1-3). However, in clinical practice NAFLD still remains an under-recognized complication of T2DM, unlike the other microvascular and macrovascular complications.

As discussed later, isolated steatosis and NASH may carry an increased risk of CVD and being the most common cause of death in patients with NAFLD, independent of other metabolic comorbidities. It is important for endocrinologists and primary care physicians to recognize that NAFLD in T2DM has been shown to be associated with adverse metabolic changes resulting in increased atherosclerotic disease and cardiovascular consequences (4,5).

NAFLD: KEY CONCEPTS

The incidence of NAFLD is rising, paralleling that of obesity and diabetes mellitus. There has been extensive research in the area of NAFLD, especially over the past two decades. However, given the lack of highly reliable noninvasive diagnostic methods, the burden of NAFLD probably remains overlooked. By liver ultrasound, studies have demonstrated the prevalence of NAFLD to be 24% in United States, whereas using blood testing alone this is underestimated at just 13% (6). By the gold-standard magnetic resonance imaging and spectroscopy (¹H-MRS), the prevalence of NAFLD in the general population is estimated to be 34% (7).

Unfortunately, imaging techniques cannot adequately evaluate for hepatocellular necrosis or inflammation (i.e. NASH). Studies that have utilized a liver biopsy to confirm the diagnosis of NAFLD have shown that 59% of patients with NAFLD have NASH, this being much higher in obese individuals (6). Moreover, recent studies have reported that about 18% of unselected patients with T2DM have moderate-to-severe (F2-F4) fibrosis (6,8).

NAFLD often progresses to steatohepatitis (NASH), especially in patients with T2DM. NASH is hallmarked by hepatocellular necrosis, lobular inflammation and often fibrosis. Many studies have now documented that patients with NASH and fibrosis have the worst mortality (9). As fibrosis progresses, cirrhosis develops. This rate of progression to cirrhosis is highly variable and dependent on age, BMI, blood pressure control, presence of T2DM, and degree of steatohepatitis (10). The three most relevant risk factors are obesity (excessive BMI or visceral obesity), T2DM, and presence of moderate to severe fibrosis (11). However, given the high heterogeneity in disease progression one must admit that the precise factors leading to cirrhosis remain unclear.

NASH is currently the second most common indication for liver transplantation, after hepatitis C. It is predicted to be the leading indication for liver transplantation in the next decade given the rise in incidence (8). The annual incidence of HCC in NAFLD – related cirrhosis is about 1% (1,8,12,13). Nonalcoholic steatohepatitis related cirrhosis is currently the third leading cause for HCC, after HCV and alcohol-related liver disease. Importantly, those with NASH- related HCC that undergo liver transplantation are more likely to have a higher BMI and higher rate of T2DM (13). In one study, it has also been demonstrated that HCC can develop in NASH in the absence of cirrhosis (14).

PATHOGENESIS

Of note, the pathogenesis of NASH is poorly understood in humans. Most proposed mechanisms at the molecular level have only been observed in cell systems or animal models, but not confirmed in humans. Animal models of NASH are far from ideal in resembling human disease (69). Often treatments that are promising in animal models are in discordance with results in humans – indeed, most treatments that have resolved NASH, and even fibrosis, in mice have failed so far in large RCTs. A detailed description of the potential pathways leading to steatohepatitis exceeds the scope of this review, therefore we refer the reader to recent in-depth reviews involving a broad spectrum of mechanisms involved in the development of NASH and liver fibrosis (11,69-72). In Figure 1 (below) we propose a schematic representation of the factors and many pathways leading to NASH and fibrosis.

Figure 1:

Pathogenies of NAFLD, adapted from Cusi K (11).

PNPLA3=patatin-like phospholipase domain-containing protein 3. TM6SF2=transmembrane-6 superfamily member 2. GCKR=glucokinase regulator. HSD17B13=hydroxysteroid 17-beta dehydrogenase 13. NAFLD=non-alcoholic fatty liver disease. HDL-C=high-density lipoprotein cholesterol. LDL-C=low-density lipoprotein cholesterol. VLDL=very low-density lipoprotein. CETP=Cholesteryl ester transfer protein.

Development of Hepatocyte Injury and Steatohepatitis: Role of Mitochondrial Dysfunction

It should be emphasized that the mechanisms leading to steatohepatitis in humans remain unknown. With limited exceptions that point to subtle defects in mitochondrial function in the liver of subjects with NAFLD and/or T2DM (reviewed in ref. 71), almost all of the available information has been extrapolated from cell culture studies or animal models of NASH. It is also unclear if NASH is always heralded by isolated steatosis, and what are the drivers of disease. While there is an increasing recognition that NASH is an heterogeneous disease affecting obese and non-obese individuals, disease progression is often with associated with obesity/weight gain and T2DM. Obvious limitations in obtaining sufficient liver tissue for molecular studies, as well as ethical challenges for performing paired liver biopsies before and after a given intervention, have greatly hampered our ability to make significant progress in understanding the pathogenesis of NASH in humans. However, factors associated with overnutrition and insulin resistance likely play a role in the maladaptation of mitochondrial oxidative function that leads to inefficient oxidative flux, accumulation of lipotoxic intermediates and the progression from isolated steatosis to NASH (71,93). As mentioned above, genetic factors may also regulate lipid droplet accumulation that may exacerbate disease progression. Many other trigger factors associated with endoplasmic reticulum (ER) stress, oxidative stress and inflammasome activation have been described. However, the exact temporal relationship and sequence of events remains elusive.

Normally, there is a close regulation between beta-oxidation, hepatic tricarboxylic acid (TCA) cycle activity, ketogenesis and ATP synthesis. Normally, FFAs influx is efficiently dealt through beta-oxidation. However, in states of chronic overfeeding, beta-oxidation can over time become relatively ineffective, resulting in the accumulation of hepatocyte ceramides and DAGs (as well as acylcarnitines), as seen in states of hepatic steatosis (71,75-77). As summarized in Figure 2, the current working hypothesis in NASH is that overactive hepatic TCA cycle carries the risk of overloading the mitochondrial electron transport chain and hence promoting not only the formation of toxic metabolites but the production of reactive oxygen species (ROS) and other inflammatory mediators. In this setting, it is believed that inflammatory pathways are triggered which then lead to hepatocyte necrosis and chronic inflammation, Kupffer cell activation and recruitment, as well as hepatic stellate cell activation. This disruption of the normal equilibrium between hepatocyte and its microenvironment (i.e., in particular with Kupffer cells and hepatic stellate cells, the latter promoting fibrogenesis) seems to determine the degree of hepatocyte injury and the triggering of downstream pathways that lead to cirrhosis, as reviewed in-depth elsewhere (70). However, while many recent interventions successful in animal models have failed in humans, it is of interest that there is a correlation between successful treatment for NASH in humans (with GLP-1RA or pioglitazone [8]) with studies in vivo with such interventions that restore hepatocyte TCA function and reduce intracellular toxic lipids (94, 95), giving support to the hypothesis of increased mitochondrial FFA flux as a potential therapeutic target for patients with NASH.

Figure 2.

Hepatic Mitochondrial Oxidative Dysfunction during NASH (71). Adipose tissue insulin resistance results in increased lipolysis and higher flux of FFAs into the liver (1), resulting in high rates of hepatic triglyceride accretion (2). Initial breakdown of FFA in the liver proceeds through b-oxidation, generating two-carbon units of acetyl-CoA (3). During hepatic insulin resistance, disposal of acetylCoA units through ketogenesis undergoes an early compensatory induction in simple steatosis, but is impaired in NASH (4). In spite of FFA overload, hepatic insulin resistance and steatosis result in beta-oxidation being inefficient and incomplete as evident from accumulating levels of hepatic ceramides, DAGs, and long-chain acylcarnitine (5). However, complete oxidation of acetyl-CoA units through the mitochondrial TCA cycle continues unabated during simple steatosis and NASH (6), potentially to meet the energetic demands of maintaining high rates of gluconeogenesis (7). The chronically elevated oxidative flux through TCA cycle during NASH has the potential to uncouple hepatic TCA cycle activity from mitochondrial respiration (8) by disrupting the mitochondrial electrochemical gradient and to impair ATP synthesis (9). This mitochondrial milieu could be a chronic source of ROS generation (10) and cellular inflammation, and could be a target for therapeutic manipulations. Abbreviations: Cer, ceramides; CoA, coenzyme A; DAGs, diacylglycerols; FFAs, free fatty acid; NASH, nonalcoholicsteatohepatitis; PEP, phosphoenolpyruvate; ROS, reactive oxygen species; TCA, tricarboxylic acid.

However, linking NASH only to altered mitochondrial flux is obviously an oversimplification of a complex web of many factors at play. Other pathways that have been implicated in hepatocyte injury and the development of NASH, although rather broadly, include cholesterol accumulation in hepatocytes (96) and a tangled web involving activation of apoptotic pathways with ER stress and abnormal unfolded protein response (97), as well as defects in autophagy (98). Recently, inflammasome activation has gained attention as it integrates many cytoplasmic signals into danger-associated molecular patterns (DAMPs) from diverse sources such as intracellular lipids to the gut microbiome (97, 98).

Diet and gut microbiota have been repeatedly implicated to play a role in the pathogenesis of NAFLD. In particular, fructose appears to play a role in NASH by stimulating DNL and suppressing β-oxidation of FFAs, hence leading to hepatocyte injury (99). Many studies have shown that excess fructose consumption, usually as sugar-sweetened beverages with sucrose (converted to fructose and glucose after ingestion), is associated with development of NAFLD and NASH. Obesity is also associated with a change in gut microbiota that produce more reactive oxygen species and are involved in triggering a variety of inflammatory pathways (100). However, the causative role of the gut microbiome in the development of T2DM or NAFLD remains overall poorly understood (101).

DIAGNOSIS

Having T2DM is associated with a much greater risk of NAFLD with approximately 70% of patients with T2DM having NAFLD when MRI-based techniques are used, as well as higher risk of having more advanced forms of the disease, such as fibrosis and cirrhosis (6,8,106). Despite all the current evidence, there is lack of awareness in primary care physicians and endocrinologists to evaluate patients with prediabetes or type 2 diabetes mellitus for NAFLD. Even if suspected to have NAFLD based on clinical characteristics, there is currently a lack of further investigations being undertaken as non-invasive biomarkers of the disease and even imaging, are not as reliable as wished and not available at every clinic. The widely accepted thought by primary care physicians, as well as many endocrinologists, is to not pursue any confirmatory testing to assess for the presence or degree of fibrosis, as it is believed to seldom change their management, except to re-emphasize lifestyle modifications and weight loss. However, few healthcare providers are aware about the efficacy of lifestyle changes and some currently available pharmacological agents to revert NASH, and even fibrosis, if done early and before the development of end-stage liver disease.

Early detection and treatment of NAFLD can lead to better histological and metabolic outcomes, including CVD, and improve overall morbidity and mortality. NAFLD is a diagnosis of exclusion, so it is imperative to eliminate all other causes of liver disease (such as, alcoholic liver disease, medication induced toxicity, viral or autoimmune hepatitis, hemochromatosis, alpha 1 antitrypsin deficiency, Wilson’s disease, other) prior to the diagnosis of NAFLD. Often management may require referral to hepatology and developing multidisciplinary teams (107, 108). Once NAFLD is diagnosed, there needs to be further testing to evaluate for the presence and severity of fibrosis (8).

TREATMENT

The aim of treatment for patients with NASH is to delay or reverse the progression of fibrosis and improve NASH-related morbidity/mortality due to hepatic (cirrhosis and HCC) and extra-hepatic complications, mainly cardiovascular disease (CVD). Currently there is no pharmacotherapy approved by regulatory agencies for the treatment of NAFLD, although pioglitazone is recommended by the current guidelines as a choice for patients with or without T2DM, and vitamin E for patients without diabetes (92,124). The FDA has accepted 2 endpoints as valid ones for drug approval in clinical trials: a) Resolution of the histological findings that define NASH (necroinflammation) without worsening of fibrosis, and b) Reversal of ≥1 fibrosis stage without worsening of steatohepatitis/NASH (124,126,127). Despite the many ongoing efforts to find novel pharmacological agents the first-line of treatment will always be lifestyle modification including diet, exercise and weight loss (92,124), to combat insulin resistance and the related conditions like diabetes and obesity so closely related to NAFLD (128-130).

CONCLUSION

Endocrinologists must be aware that NAFLD is a potentially severe disease in patients with T2DM, due to both its hepatic and extrahepatic complications. In 2019 the ADA included for the first time in its recommendations to implement regular screening for advanced fibrosis in all patients with prediabetes or T2DM with evidence of elevated plasma aminotransferases or steatosis, so an early diagnosis can prevent long-term complications (118). This is the first step of management while being aware of the significant need for accurate and cost-effective diagnostic modalities and for continued research efforts for new treatments.

Figure 3 is a suggested algorithm to be used for endocrinologists and primary care settings when evaluating a patient with prediabetes or T2DM for the possibility of having NASH.

In the future, we anticipate that patients with T2DM will be routinely screened for NASH in the same way they are today for diabetic retinopathy or nephropathy.

Figure 3.

Management of patients with prediabetes or type 2 diabetes mellitus and suspected NAFLD. Based on figure from reference (8). *High risk patients include patients with type 2 diabetes > 10 years, A1c > 8.5%, triglycerides > 250mg/dl, evidence of steatosis based on MR imaging or controlled attenuation parameter (CAP), or genetic testing (PNPLA3 and/or TM6SF2).

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*

equal effort

†

equal effort